Isolation and Culture of Alveolar Type Ⅱ Epithelial Cells and Injurious Effects of Phospholipase A_2 on the Cultured Cells / 第二军医大学学报

ABSTRACT

A method for isolation and primary culture of guinea pig alveolar type Ⅱ epithelial cells is reported. The injurious effects of phospholipase A2 (PLA2) on the cultured cells were studied. Trypsinizing solution was instilled into the bronchoalveolar space. The lungs were digested, sectioned, vortexed and filtered. The yield of original cell mixture was 172.3?106 cells/guinea pig, of which 33.6% were type Ⅱcells. At 48 h of culture following short term selective attachment 7.9?106 cells were obtained, of which 66.1% were type Ⅱ ceils. The purity could further be raised to 76.2% (63.9% -92%) if pretreated with ethyienediarnine tetraacetic acid (EDTA) before trypsinization to remove attached cells. The characteristics of cultured cells were observed under phase contrast microscopy, light microscopy and electron microscopy. After exposure of the cells to 300U/ml PLA2 for 4h at 37℃, detachment and lactic dehydrogenase (LDH) release of the cultured cells increased significantly. It is suggested that PLA, may damage and promote detachment of cultured type Ⅱ cells and it may play an important part in the development of some diseases.